The transition from drug use to abuse and, eventually, to dependence may be mediated by biological factors that are present prior to drug use. Although chronic exposure to drugs of abuse is known to disrupt many signaling pathways, little is known about the molecular mechanisms that mediate addiction susceptibility. There is considerable interest in identifying early biomarkers for addiction susceptibility to improve addiction prevention strategies. Most studies have been conducted in substance-dependent individuals where the dissociation between ?susceptibility? and ?consequence? is ambiguous. I have recently identified a behavioral phenotype in rats that reliably predicts future drug-taking behaviors and identified three proteins as potential mediators of addiction susceptibility: sorting nexin 1 (SNX1), ryanodine receptor 2 (RYR2), and ataxin 2-like (ATXN2L). These proteins are involved in intracellular trafficking, calcium signaling and cytoskeleton reorganization, and have been previously linked to addiction. Precisely how differences in expression of these proteins impact the signaling cascades underlying addiction susceptibility is not known. The overarching goal of this proposal is to determine the functional and molecular role of proteins that mediate addiction susceptibility and investigate how these factors are mechanistically linked to genetic and/or environmental components of risk for addiction. To accomplish this goal, the proposed research will combine sophisticated behavioral and computational assessments with viral, proteomic and bioinformatic approaches. In Aim 1 I will use an inducible and reversible viral construct to bi-directionally manipulate expression of SNX1, RYR2, and ATXN2L and also determine how changes in expression of SNX1, RYR2, and ATXN2L alter methamphetamine self-administration and protein signaling mechanisms. In Aim 2 I will determine if variation in the expression of SNX1, RYR2, and ATXN2L is altered in a model of genetic addiction susceptibility and is associated with increased addiction risk. In Aim 3 I will determine if variation in expression of SNX1, RYR2, and ATXN2L is altered in a model of environmental addiction susceptibility and is associated with increased addiction risk. Completion of these aims will generate new insights into the signaling mechanisms of addiction susceptibility that could identify early biological markers of risk for addiction and improve current strategies for addiction prevention. The Principal Investigator will receive mentorship and technical training in viral and proteomic technologies by experts in cell signaling and cellular mechanisms, and viral technologies in motivated behaviors. Yale University and the Department Psychiatry provide exceptional facilities and resources for completing the proposed experiments, as well as having an exceptional reputation and track record for mentoring and transitioning early-stage investigators in to independent investigators. The proposed training, education and research will provide the PI with the technical and professional training to become a successful, independent addiction investigator.